Improving the Application of Prothrombin Time-Derived Fibrinogen Assay Through Value Transfer from the Von Clauss Method.

BACKGROUND: This study aimed to improve the accuracy of the fibrinogen (Fib) prothrombin time-derived (PT-der) method. To achieve this, a value transfer method was introduced for calibration, and its effectiveness was assessed. METHODS: The PT-der Fib assay was calibrated by pooled samples (assigned by the von Clauss method) in three different ways: 1) multipoint calibration using an automatic dilution system, 2) multipoint calibration using a manual dilution method, and 3) manual calibration with multiple concentrations. Three calibration equations (1, 2, and 3) were obtained and an optimal equation was selected by comparing the detection results of the von Clauss method with the PT-der method. Subsequently, the optimal equation was assessed for an accuracy limit, and linear analysis and reference interval verification were performed following the guidelines (EP15-A and EP6-A) issued by the CLSI. RESULTS: Compared with the other two equations (equation 1 and 2), equation 3, available from manual calibration with multiple concentrations, showed a better performance for the PT-der determination in a primary cohort (n = 208), and a good agreement (99% of the results between 1.52 and 6.30 g/L were interchangeable) was validated (n = 3226). The reference interval was also verified in almost all healthy individuals (39/40). However, the discrep-ancy between the two methods was observed in several specific conditions, such as hyperfibrinolysis. CONCLUSIONS: Manual calibration with multiple concentrations is better for the Fib PT-der method assay. As a rapid, accurate, and economical test, the performance of the Fib PT-der method has been verified and may be more applicable than before.

A high throughput metabolomics method and its application in female serum samples in a normal menstrual cycle based on liquid chromatography-mass spectrometry.

Periodical changes of steroid hormones have a great impact on the homeostasis of the female. However, there are few studies concerning the metabolome changes during the cycle. To study the periodic metabolic changes, a female cohort was enrolled with time-series serum samples collected during a menstrual cycle. To meet the requirement of the large-scale sample analysis, a high throughput metabolomics method was established by using an efficient sample preparation on a 96 well filter plate and a rapid LC condition in 12 min, which reduces about 70% of the samples preprocessing time and 60% analysis time. Evaluation of metabolite coverage and separation performances reflected that the method was robust for the large-scale metabolomics study. Using this method, we found that 12.6% of total detected ions including lipids, amino acids, citric acid, and so on were significantly changed during a menstrual cycle. Some metabolites were found periodically changed, which is similar to hormones (estrone and progesterone) during the cycle. These results show the novel high throughput method can be applied in large-scale metabolomics studies.

Effect of traditional Chinese medicinal enemas on ulcerative colitis of rats.

AIM: To investigate the effects of traditional Chinese medicinal enema (TCME) on inflammatory and immune response of colonic mucosa of rats with ulcerative colitis (UC), and to observe the pathogenic mechanism. METHODS: Thirty UC rats, induced by intestinal enema together with 2.4-dinitrochlorobenzene (DNCB) and acetic acid, were randomly divided into 3 groups, i.e., GI, GII and GIII. Groups GI and GII were administered with TCME and salazosulfapyridine enema (SASPE), respectively. Group GIII was clystered with only normal saline (NSE), served as control. Group GIV was taken from normal rats as reference, once daily, from the 7th day after the establishment of UC for total 28 d. Interleukin-6 (IL-6) in the colonic mucosa was assayed by (3)H-TdR incorporation assay. Colonic mucosal lymphocyte subpopulation adhesive molecules, CD(4)(+)CD(11a )(+), CD(4)(+)CD(18)(+), CD(8)(+)CD(11a)(+), CD(8)(+)CD1(18)(+) (LSAM), tumor necrosis factor (TNF)-alpha, and interferon-gamma (IFN-gamma), were detected by enzyme linked immunosorbent assay (ELISA). Moreover, the expression of TNF-alpha mRNA and IFN-gamma mRNA in colonic mucosa were detected by polymerase chain reaction (RT-PCR). RESULTS: Before therapies, in model groups, GI, GII and GIII, levels of IL-6, TNF-alpha, IFN-gamma, CD(8)(+)CD(11a)(+) and CD(8)(+)CD(18)(+) were significantly different (38.29+/-2.61 U/mL, 16.54+/-1.23 ng/L, 8.61+/-0.89 ng/L, 13.51+/-2.31% and 12.22+/-1.13%, respectively) compared to those in GIV group (31.56+/-2.47 U/mL, 12.81+/-1.38 ng/L, 5.28+/-0.56 ng/L, 16.68+/-1.41% and 16.79+/-1.11%, respectively). After therapeutic enemas, in GI group, the contents of IL-6 (32.48+/-2.53 U/m), TNF-alpha (13.42+/-1.57 ng/L) and IFN-gamma (5.87+/-0.84 ng/L) were reduced; then, the contents of CD(8)(+)CD(11a)(+) (16.01+/-1.05 %) and CD(8)(+)CD(18)(+) (16.28+/-0.19%) were raised. There was no significant difference between groups GI and GIV, but the difference between groups GI and GII was quite obvious (P<0.05). The expressions of TNF-alpha mRNA and IFN-gamma mRNA in group GIII were much higher than those of group GIV, but those in group GI were significantly suppressed by TCME therapy. CONCLUSION: Ulcerative colitis is related to colonic regional mucosal inflammatory factors and immune imbalance. TCME can effectively inhibit regional mucosal inflammatory factors and improve their disorder of immunity.

[Exploration of toxicity reducing mechanism of aconite alkaloids during decoction process using liquid chromatography-mass spectrometry].

A high performance liquid chromatography-electrospray ionization mass spectrometry method was developed for the determination of aconite alkaloids. It was used to investigate the degradation of alkaloids of Radix Aconiti Lateralis Preparata during decoction. Six alkaline degradation products were identified, and the degradation regularity of diester-diterpenoid alkaloids was confirmed during the test using standards. The dynamic changes of the amount of aconite alkaloids in the decoction of Radix Aconiti Lateralis Preparata were supervised. Along with the increase of decoction time, the concentrations of diester-diterpenoid alkaloids and lipo-alkaloid decreased significantly. The results can provide a scientific basis for the safety use of aconite.